D zone test: Introduction, test procedure and clinical significance

D Zone Test: Introduction, Test Procedure and Clinical Significance

Introduction of D-Zone Test

This test is most important in Staphylococcus aureus. If the D zone test is positive, the organism is iMLSB strain. The test is called so because of the zone of inhibition like the D zone and the antimicrobial drug’s uses are erythromycin and clindamycin. The above figure is showing D-zone test positive i.e. Macrolides, lincosamides, and type B streptogramins (iMLSB resistance) strain.

Mechanism of D Zone Test ( iMLSB strain)

Erythromycin ((a macrolide) and clindamycin (a lincosamide) represent two distinct classes of antimicrobial agents that inhibit protein synthesis by binding to the 50S ribosomal subunits of bacterial cells. In staphylococci, resistance to both of these antimicrobial agents can occur through methylation of their ribosomal target site. Such resistance is typically mediated by erm genes. Resistance to macrolides also can occur by efflux, typically mediated by the msrA gene. Another resistance mechanism, the inactivation of lincosamide by chemical modification (such as mediated by the inuA gene), appears to be rare. The target site modification mechanism, also called Macrolide-Lincosamide-Streptogramin B (MLSB) resistance, results in resistance to erythromycin, clindamycin, and streptogramin B. This mechanism can be constitutive, where the rRNA methylase is always produced, or can be inducible, where methylase is produced only in the presence of an inducing agent. Erythromycin is an effective inducer, but clindamycin is a weak inducer. In vitro, Staphylococcus aureus isolates with constitutive resistance are resistant to erythromycin and clindamycin, and isolates with inducible resistance are resistant to erythromycin but appear susceptible to clindamycin. In vivo, therapy with clindamycin may select for constitutive erm mutants, which may lead to clinical failure Isolates with msrA-mediated efflux also appear erythromycin-resistant and clindamycin susceptible by in vitro tests; however, such isolates do not typically become clindamycin resistant during therapy. An in vitro induction test can distinguish staphylococci that have inducible erm-mediated resistance from those with msrA mediated resistance.

D zone Test Procedure

The test is performed by disk diffusion, placing a 15-µg erythromycin disk in proximity to a 2-µg clindamycin disk on an agar plate that has been inoculated with a staphylococcal isolate; the plate is then incubated overnight. A flattening of the zone of inhibition around the clindamycin disk proximal to the erythromycin disk (producing a zone of inhibition shaped like the letter D) is considered a positive result and indicates that the erythromycin has induced clindamycin resistance (a positive “D-zone test”). For erythromycin-resistant isolates, induction tests can help laboratories determine whether results for clindamycin should be reported as susceptible (when the induction test is negative) or as resistant (when the induction test is positive).

Clinical significance of D zone Test

D-zone test positive organisms i.e. i MLSB strain are resistant to three groups of antimicrobial drugs like macrolide, lincosamide, and streptogramin B.

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